ABSTRACT Gamma-hydroxybutyrate (GHB), is a popular drug of abuse utilized at raves and in drug-facilitate sexual assault due to its? euphoric, aphrodisiac, and sedative effects. The annual number of GHB-associated deaths has continued to increase since the 1990s in the United States, the United Kingdom, Western Europe and Australia. In the last decade the use of GHB has been increasing in the LBGTQ community due to the prevalence of a phenomenon referred to as chemsex. Of the drugs used for chemsex, GHB is the most likely to cause acute overdose. Proton- and sodium-dependent monocarboxylate transporters (MCTs/SMCTs) are involved in the transport of GHB across biologically important barriers and tissues, and their expression governs GHB renal clearance and brain distribution. Preliminary data demonstrates that GHB toxicokinetics (plasma concentrations and renal clearance) are altered in the presence and absence of female sex hormones. Further, we have demonstrated that male and female sex hormones regulate monocarboxylate transporters in the kidney, and differences in GHB renal clearance over the estrous cycle, and between males and females are consistent with the changes in renal monocarboxylate transporter expression. Despite the potential for sex hormones to regulate GHB renal clearance and tissue distribution, there is a paucity of information in the literature regarding the influence of sex and cross-sex hormone treatment on GHB toxicokinetics and toxicity. The focus of this application is on investigating GHB toxicokinetics in response to sex and cross-sex hormone therapy and identifying the sex hormone-dependent mechanisms regulating expression of renal and blood brain barrier (BBB) monocarboxylate transporters. Our overall hypothesis is that variability in GHB toxicokinetics and overdose risk result from sex hormone-dependent regulation of monocarboxylate transporters that govern GHB clearance and distribution. We have proposed two specific aims to evaluate this hypothesis utilizing in vivo models of sex and cross-sex hormone replacement, molecular biological and toxicokinetic techniques. The first specific aim investigates GHB toxicokinetics and toxicity in response to sex and cross-sex hormone treatment. Our hypotheses for this aim are that [1] GHB renal clearance and systemic exposure (AUC) will be altered in response to individual male and female sex hormones; [2] males, and animals exposed to testosterone will have an increased risk of acute overdose due to decreased renal clearance. In our second aim, we will investigate the sex hormone-dependent regulation of renal and BBB MCTs and SMCTs. Our hypotheses are that sex and cross- sex hormone treatment will differently regulate monocarboxylate transporter expression, and this regulation is sex hormone receptor dependent. This proposal represents a novel extension of our previous work, and will further our mechanistic understanding of sex hormone-dependent regulation of drug clearance, and the resultant toxicokinetic consequences, and will help to identify populations with a greater risk of GHB overdose.